Encephalitis and meningitis are potentially fatal diseases defined by acute inflammation of the brain or protective membranes covering the brain and spinal cord. These diseases can be caused by viruses, bacteria, fungi, parasites or amoeba, and disproportionately affect children, the elderly, or the immunocompromised. Viruses are the most common cause of encephalitis, with the majority of aseptic encephalitis cases caused by enterovirus, human herpesviruses, and arboviruses. Unfortunately, clinical presentation and initial laboratory findings in cases of meningitis and encephalitis are usually too nonspecific to permit an etiologic diagnosis. Complications arising from central nervous system (CNS) infection and appropriate treatment strategies depend on the organism involved. It is therefore important to differentiate between those cases for which a specific treatment has been shown effective, and those for which only supportive treatment is indicated. Nucleic acid amplification and detection assays have been considered the test of choice for CNS infections for more than a decade, yet there are only two FDA-approved molecular diagnostic tests for use with cerebrospinal fluid (CSF), each of which only detects enterovirus. Given the invasiveness of lumbar puncture, the need for low limits of detection in CSF, the range of organisms causing aseptic meningitis or encephalitis, the clinical advantage of detection in facilitating appropriate treatment, the many practical advantages of achieving multiple test results on a single small sample volume, the lack of FDA-cleared tests for diagnosis, and the importance of global surveillance activity in disease control and prevention, it is important to develop a sensitive, multiplexed diagnostic test for these diseases. The assay developed in Phase 1 was derived from a set of real-time PCR and reverse transcriptase PCR assays developed by the Laboratory of Viral Diseases at the Wadsworth Center, Akonni single-chamber amplification microarrays, and a low-cost instrument package. Viruses are detectable at d 100 gene copies per reaction, meeting the Phase 1 sensitivity milestone. The objectives of Phase 2 are to complete a sample- to-answer system, evaluate its clinical performance, and prepare for commercial production in compliance with the 21 CFR 820, the FDA's Quality System Regulation.